Human hematopoietic stem cells (HSCs) transplants can treat a range of hematological malignancies and genetic blood disorders. However, success is limited by a lack of optimal donors and low number of stem cells available from common HSC sources. To date, expansion of HSCs ex vivo for enhanced in vivo engraftment in patients has been clinically ineffective. Insufficient cell numbers generated in culture or poor differentiation of the starting cell population ex vivo has been contributing factors to improper clinical biomanufacturing of these cells. The current techniques of hematopoietic cell biomanufacturing are expensive which further complicates scalability and wide clinical translation. To overcome these critical barriers and help achieve the full life- saving potential of HSCs, novel approaches to maintain and expand patient-derived HSCs in vitro are needed. Stem cell self-renewal and differentiation are regulated through intricate crosstalk with neighboring cell types, which secrete and organize a multifaceted milieu of signaling cues (stem cell niche). Removing stem cells from their native environment can disrupt this homeostasis. HSCs experience limited self-renewal in the bone marrow niche (BM) and are typically quiescent. In contrast, in fetal liver, HSCs undergo marked expansion and become highly proliferative, which suggests that the fetal liver niche provides a unique microenvironment for HSCs. However, access to viable human fetal liver is challenging due to ethical constraints. By genetically engineering human induced pluripotent stem cells (hiPSCs), for the first time we could generate a fetal liver tissue with hematopoietic niche capacity. In our approach, a transient and heterogeneous pulse of GATA6 transcription factor for 5 days resulted in co-development of mesoderm and endoderm layers in culture. The culture further self-organized into a functional human fetal liver tissue (containing hematopoietic cells) without the need to add exogenous growth factors to the culture. Our objective is to develop a universal, and common platform for expansion of human HSCs that is scalable, simple, and economical. We hypothesize that our human fetal liver tissue autonomously produces known and unknown factors that contribute to hematopoiesis and can provide us with a ?universal? and ?programmable? cellular microenvironment, or niche for this purpose. In aim 1 we will employ a GATA6-engineered Fetal LIver Niche (FLIN) for the expansion of HSCs. In aim 2, we will interrogate hematopoietic niche environment through engineering a customizable fetal liver, DESigner Liver Niche (DESLIN) and in aim 3, we will examine scalability of FLIN-HSC cultures in microcarrier-based Stirred suspension bioreactors. Overall, the development and optimization of this platform has the potential to dramatically reduce the cost of large-scale production of HSCs and will shed light on the biology and key signaling molecules affecting hematopoiesis. Subsequently, the delivered knowledge and tools will be applicable in a broad spectrum of hematological diseases including malignancies and genetic disorders.